Beta-2-microglobulin levels in the cerebrospinal fluid of normal dogs and dogs with neurological disease

BACKGROUND: Cerebrospinal fluid (CSF) analysis is the basis for establishing a diagnosis of central nervous system (CNS) inflammation. However, the information provided by routine CSF analysis is limited. Determination of CSF beta-2-microglobulin (beta2m) concentration has been used diagnostically in humans to identify inflammatory CNS disease; we hypothesized that it may have similar value in dogs. OBJECTIVES: The objective of this study was to measure (beta2m concentration in the CSF of clinically healthy dogs and compare the values to those observed in dogs with inflammatory CNS disease and intervertebral disc disease (IVDD). METHODS: CSF was collected from 10 clinically healthy laboratory dogs and 11 dogs each with inflammatory CNS disease and IVDD. Routine CSF analysis was performed, and (beta2m concentration was measured by ELISA. CSF (beta2m concentration and CSF:serum (beta2m ratio were compared between groups by ANOVA. Linear relationships between CSF total nucleated cell count (TNCC), RBC count, total protein concentration, and (beta2m concentration were assessed by regression analysis. RESULTS: The mean (+/- SD) CSF (beta2m concentration in clinically healthy dogs was 0.36 (+/- 0.05 microg/mL (cisternal) and 0.40 (+/- 0.07 microg/mL (lumbar). Median CSF (beta2m concentration in dogs with IVDD (0.46 microg/mL) and inflammatory CNS disease (0.85 microg/mL) differed from that of controls (0.36 microg/mL; P=.002). The concentration also differed between the 2 disease groups (P=.01). Five dogs with inflammatory CNS disease had CSF:serum (beta2m ratios >1. A correlation was identified between TNCC and (beta2m concentration (r=0.69, P=.0003). CONCLUSIONS: CSF (beta2m concentration is higher in dogs with IVDD and inflammatory CNS disease, with highest values seen with inflammatory disease. This may be attributed in part to the correlation between CSF (beta2m concentration and TNCC, but also may reflect intrathecal immune activation.     

Evaluation of the ADVIA 120 for analysis of canine cerebrospinal fluid

BACKGROUND: Conventional techniques for canine cerebrospinal fluid (CSF) analysis require large sample volumes and are labor intensive and subject to operator variability. Objective: The purpose of this study was to evaluate the ADVIA120 CSF assay for analysis of canine CSF samples. METHODS: CSF samples collected from 36 healthy control dogs and 17 dogs with neurologic disease were processed in parallel using the automated assay and established manual methods using a hemocytometer and cytocentrifugation. Results for WBC (total nucleated cell) count, RBC count, and differential nucleated cell percentages were compared using Spearman rank correlation coefficients and Bland-Altman bias plots. RESULTS: Correlation coefficients for WBC and RBC counts were 0.57 and 0.83 for controls, and 0.92 and 0.94 for ill dogs, respectively. Coefficients for the percentages of neutrophils, lymphocytes, and monocytes were 0.53, 0.26, and 0.12 for controls and 0.77, 0.92, and 0.70 for dogs with neurologic disease. When data were combined (n=53), correlation coefficients were 0.86 and 0.91 for WBC and RBC counts, and 0.63, 0.43, and 0.30 for neutrophil, lymphocyte, and monocyte percentages. A 9.5% positive bias and 7.0% negative bias were obtained for the ADVIA 120 CSF assay for lymphocytes and macrophages in dogs with neurologic disease with Bland-Altman analysis. A 12.2% positive bias was found for lymphocyte percentage in dogs with neurologic disease. CONCLUSIONS: Manual and automated CSF assays had moderate to excellent correlation for WBC and RBC concentrations, but results were more variable for differential cell percentages. The ADVIA assay may be more useful for assessment of canine CSF with adjustment of cell differentiation algorithms.     

Cytologic interpretation of canine cerebrospinal fluid samples with low total nucleated cell concentration,with and without blood contamination

Background: Cerebrospinal fluid (CSF) is potentially altered by iatrogenic blood contamination at the time of sampling due to the addition of blood‐associated leukocytes and protein. Objectives: The objective of this study was to assess whether protein concentration, neutrophil percentage, and the presence of activated macrophages, reactive lymphocytes, or eosinophils in CSF samples with low total nucleated cell concentration (TNCC) are affected by blood contamination or associated with central nervous system (CNS) disease. Methods: Case records from the Royal Veterinary College Diagnostic Laboratory were searched retrospectively for dogs with CSF having ≤5 TNCC/μL. TNCC, RBC, and protein concentrations; neutrophil percentage; and the presence of activated macrophages, reactive lymphocytes, and eosinophils were recorded. Results of magnetic resonance imaging (MRI) also were recorded as a marker of CNS disease. Results: Of 906 cases evaluated, 106 (12%) had blood contamination (>500 RBCs/μL) in CSF. Protein concentration and neutrophil percentage were significantly higher and the presence of eosinophils was more likely in blood contaminated vs noncontaminated samples. Non‐blood‐contaminated samples with activated macrophages or reactive lymphocytes had higher protein concentrations and neutrophil percentages, and those with activated macrophages were more likely to have a positive finding on MRI. Conclusions: Protein concentration, neutrophil percentage, and the presence of eosinophils are significantly affected by blood contamination in canine CSF having low TNCC. Activated macrophages and reactive lymphocytes are not affected by blood contamination, however, and may be useful in identifying dogs with CNS abnormalities.     

Fungal infections of the central nervous system in the dog and cat

Fungal infections of the central nervous system (CNS) in dogs and cats are uncommon. The purpose of this paper is to review the clinical signs, diagnostic tests, and therapeutic options of fungal infections of the CNS in the dog and cat. Clinical signs are dependent on lesion location and are often multifocal. Extraneural involvement is common. Antemortem diagnosis can be difficult and is definitively made via cytology, biopsy, or culture of an affected organ or cerebrospinal fluid (CSF). Magnetic resonance imaging can support a diagnosis and may assist in therapeutic decisions. Fungal serology can support a diagnosis when direct visualization of the organism is not possible. Long-term azole maintenance therapy is suggested to enhance survival and prevent relapse. Serial cerebrospinal fluid evaluation and magnetic resonance imaging may identify early relapse.     

Prevalence and significance of extracellular myelin‐like material in canine cerebrospinal fluid

Background: Myelin‐like material in canine cerebrospinal fluid (CSF) specimens has been attributed to demyelinating or myelomalacic conditions. In our experience, myelin‐like material is observed frequently, especially in lumbar samples, and in a variety of disease conditions. Objectives: The objective of this study was to determine if there are associations between the presence of myelin‐like material and CSF collection site, body weight, underlying disease, and patient outcome. Methods: Wright–Giemsa‐stained cytocentrifuged specimens of CSF from the cerebellomedullary cistern (n=51) and lumbar cistern (n=47) of 98 dogs with neurologic disease were evaluated retrospectively for the presence and amount of extracellular myelin‐like material. Results were compared based on collection site, body weight, type of neurologic disease, and outcome. Results: Myelin‐like material was observed in 20/98 (20%) samples and was more frequently observed in lumbar (17/47, 36%) than cerebellomedullary samples (3/51, 6%) (P=.0028). Samples from dogs <10 kg were more likely to contain myelin (14/36, 39%) compared with dogs ≥10 kg (5/60, 8%) (P=.0052). Larger amounts of myelin‐like material were observed in CSF from dogs with intervertebral disk disease compared with other diseases (P=.045). No association was found between myelin‐like material and outcome. Conclusion: The association of extracellular myelin‐like material in canine CSF samples with sampling site and body weight suggests it is more often an artifact of collection technique and anatomy rather than the result of neurologic disease. Myelin‐like material in CSF is not associated with a poorer prognosis.     

The Clinical Significance of Cerebrospinal Fluid Lipid Peroxides in Central Nervous System Disease

Forty-four dogs were referred to our hospital presenting with neurological symptoms such as seizure or paraparesis. Magnetic resonance imaging (MRI) revealed abnormal results in 21 (abnormal MRI group) and normal results in 23 dogs (normal MRI group). Cerebrospinal fluid (CSF) (normal MRI group, n = 22; abnormal MRI group, n = 21) and serum lipid peroxide (LP) concentrations (normal MRI group, n = 11; abnormal MRI group, n = 15) were measured in a number of these dogs, and revealed a significant difference in the CSF/serum LP values (normal MRI group, n = 10; abnormal MRI group, n = 14) between the abnormal and the normal MRI groups (t-test and Mann–Whitney U-test p < 0.05). No other significant differences were observed. CSF/serum LP values exceeding 1.0 were exhibited in 10 of 14 dogs (71%) in the abnormal MRI group, and in 1 of 10 dogs (10%) in the normal MRI group. In the remaining animals, 4 dogs of the abnormal MRI group showed CSF/serum values lower than 1.0, 3 dogs had morphological abnormalities but no abnormal MRI signals in the central nervous system, and 1 dog had an abnormal MRI signal but no pathological abnormality. In the CSF analysis, 3 of 16 dogs (19%) of the abnormal MRI groupshowed abnormal cell counts and/or protein content. We conclude that the CSF/serum LP value can be used for the detection of neurological lesions such as oedema, inflammation and tumour.     

Clinical, cerebrospinal fluid, and histological data from thirty-four cats with primary noninflammatory disease of the central nervous system.

The purpose of this study was to report the clinical, cerebrospinal fluid (CSF), and histological data derived from a study of 34 cats with noninflammatory central nervous system (CNS) disease, and to report the activities of the enzymes lactate-dehydrogenase (LDH), aspartate transferase (AST), and creatine kinase (CK) in the CSF from 15 cats with a variety of CNS diseases. The cats were part of a study of 61 cats that were admitted to two university clinics because of signs of CNS disease. The most frequent noninflammatory diseases were neoplasia (n = 12) and ischemic encephalopathy (n = 4). The majority of cats with CNS neoplasia had a mild increase in CSF protein concentration (less than 1 g/L [100 mg/dL]), an increased percentage of neutrophils or lymphocytes, and a normal total white cell count. Cats with ischemic encephalopathy (IE) had a mild to moderate increase in CSF protein concentration (< or = 2 g/L [200 mg/dL]) and a mild increase in white cell count (< or = 10 cells/microL) with an increased percentage of lymphocytes. The enzymes LDH, AST, and CK in the CSF were not sensitive indicators of chronic CNS disease. The CSF differential cell count was frequently abnormal when the total white cell count was normal, and blood contamination in the CSF samples was a frequent problem that had to be considered in the interpretation of the results. The history, signalment, and clinical signs, when combined with the CSF findings, were valuable in the diagnosis of noninflammatory CNS disease.     

Clinical, cerebrospinal fluid, and histological data from twenty-seven cats with primary inflammatory disease of the central nervous system.

The purpose of this report is to present the clinical, cerebrospinal fluid (CSF) and histological data from 27 cats with inflammatory disease of the central nervous system (CNS). The cats were part of a study of 61 cats admitted to two university clinics over an eight-year period because of signs of CNS disease. The most frequent diseases were feline infectious peritonitis (FIP) (12/27) and suspected viral disease other than FIP (10/27). Typical CSF findings in cats with FIP were a protein concentration of greater than 2 g/L (200 mg/dL) and a white cell count of over 100 cells/microL, which consisted predominantly of neutrophils. In contrast, the CSF of cats with suspected viral disease had a protein concentration of less than 1 g/L (100 mg/dL) and a total white cell count of less than 50 cells/microL. In general, cats with FIP or suspected viral disease were less than four years of age. Neurological signs were usually multifocal in cats with FIP, but focal in cats with suspected viral disease. The CSF findings were variable in five other inflammatory diseases represented. Two cats with protozoan infection had normal CSF total cell counts but abnormal differential counts. The CSF findings were invaluable in differentiating FIP from other causes of inflammatory CNS disease.     

Diverse functions of perlecan in central nervous system cells in vitro

Therapeutic treatment targeting one cell type is considered ineffective in remedying any injury to the central nervous system (CNS). Perlecan, a multi‐functional, heparan sulfate proteoglycan, shows diverse effects on distinct cell types, suggesting that it is one of the candidates that can augment the regenerative mechanisms in the injured CNS. Therefore, we examined the functions of perlecan in CNS cells in vitro by using perlecan purified from bovine kidney. Perlecan‐coated cell culture plates, unlike their type I/III collagen‐coated counterparts, did not inhibit the adhesion of neural stem/progenitor cells (NS/PCs) and neurons. The coated perlecan and the perlecan added to the culture medium suppressed astrocyte proliferation; however, perlecan added to the medium promoted NS/PC proliferation. Neurons were promoted to extend their neurites on the perlecan‐coated substrate, and perlecan added to the medium also showed a similar effect. NS/PC proliferation and neurite extension is a major regenerative reaction in CNS injury, whereas excess proliferation of astrocytes cause hypertrophy of glial scars, which repels neurons. Our in vitro study suggests that perlecan is an attractive candidate to promote regenerative mechanisms and to suppress reactions that hamper regenerative processes in cases of CNS injury.     

Total protein, albumin quota, and electrophoretic patterns in cerebrospinal fluid of dogs with central nervous system disorders

Cerebrospinal fluid was analyzed for total protein, albumin quota, and electrophoretic patterns of albumin and alpha-, beta-, and gamma-globulins in 10 healthy dogs and 35 dogs with CNS disorders. Values obtained in healthy dogs were used to establish control data. Samples also were collected from dogs with neurologic diseases that were classified according to clinical and pathologic diagnosis as inflammation, neoplasm, and spinal cord compression. The albumin quota and total CSF albumin values were used as indicators of blood-brain barrier disturbance. Four patterns were observed on agarose electrophoresis that included intrathecal immunoglobulin production, intrathecal immunoglobulin production combined with blood-brain barrier disturbance, blood-brain barrier disturbance, and unaltered CSF. These patterns correlated with the observed clinical and pathologic conditions. Seemingly, agarose electrophoresis of CSF is a simple and reliable technique that aids in the diagnosis of CNS disorders of dogs.     

Effects of time, initial composition, and stabilizing agents on the results of canine cerebrospinal fluid analysis

BACKGROUND: Cerebrospinal fluid (CSF) is considered highly labile, but not all samples are analyzed immediately. Changes in the composition of CSF could potentially affect diagnostic test results and thus influence decisions about patient management. There has been little scientific inquiry into how variables such as time, initial composition, and storage conditions affect results of standard laboratory analysis of CSF. OBJECTIVES: The objectives of this study were to determine the effects of time, protein concentration, and presence or absence of exogenous stabilizing agents on standard CSF analysis results. METHODS: Thirty abnormal CSF samples from 26 dogs were evaluated. Samples were divided into aliquots comprising different treatment groups and stored at 4 degrees C. Total nucleated cell count (TNCC), differential cell count (DCC), and cell morphology were evaluated for all groups; protein concentration was measured for selected groups. Unaltered aliquots were analyzed immediately (T0Hr) and at 2, 4, 8, 12, 24, and 48 hours (T2Hr-T48Hr); aliquots with added fetal calf serum (FCS) or hydroxyethyl starch (hetastarch) were analyzed at T48Hr. RESULTS: Significant time-dependent changes were observed in DCC in unaltered samples. Mononuclear cells deteriorated more rapidly than did neutrophils. Based on microscopic examination and subjective scoring of cell morphology, cells were consistently more degenerate by T24Hr compared with T0Hr. Samples with protein concentrations > or =50 mg/dL were less susceptible to cell deterioration than those with lower protein concentrations. Adding either FCS or hetastarch improved sample stability. CONCLUSIONS: Delayed analysis of canine CSF by 4-8 hours is unlikely to alter diagnostic interpretation, especially for samples with protein concentrations > or =50 mg/dL. The likelihood of misinterpretation is higher for samples with low cellularity or low protein concentration. We provide specific recommendations for adding FCS or hetastarch to samples that will not be analyzed within 1 hour.     

Establishment and characterization of a canine sebaceous epithelial cell line derived from an eyelid mass

Little is known about the pathological roles of sebaceous glands in canine skin diseases, as most examinations have been conducted with cultured human sebaceous epithelial cell lines. To our knowledge, there is no available canine sebaceous epithelial cell line. The purpose of this study was to establish a canine sebaceous epithelial cell line and characterize it. An eyelid mass in a dog was surgically resected for treatment, and it was histologically diagnosed as sebaceous epithelioma. Collected tissue was conducted for culture, and the growing epithelial-like cells were passaged. The cells showed continuous proliferation for over 6 months. After 40 passages, the cells were named CMG-1. Lipid droplets in the cytoplasm of CMG-1 cells were confirmed by Oil Red O staining. As reported in studies with human sebaceous epithelial cell lines, lipogenesis in CMG-1 cells was promoted by linoleic acid, whereas transforming growth factor-β (TGF-β) suppressed it. Additionally, real-time PCR revealed that the expression levels of chemokines and cytokines, including CC chemokine ligand (CCL)-2, CCL-20, CXCL-10, Tumor necrosis factor-α (TNF-α), Interleukin (IL)-1α, IL-1β, and IL-8, were significantly increased in CMG-1 cells following treatment with lipopolysaccharide. In conclusion, we successfully established a new canine sebaceous epithelial cell line. Our data indicated that lipogenesis and inflammatory responses were quantitatively evaluable in this cell line. CMG-1 cells could be useful for the pathological analysis of sebaceous gland diseases in dogs.     

Bartonella species antibodies and DNA in cerebral spinal fluid of cats with central nervous system disease

Bartonella species infection is associated with central nervous system (CNS) disease in some humans and cats but the diagnosis is difficult to confirm with blood or serum test results. In this retrospective study of 100 client-owned cats, serum and cerebral spinal fluid (CSF) were assayed for Bartonella species IgG antibodies and CSF was assayed for Bartonella species DNA. Bartonella species IgG antibodies were detected in serum of 36 cats, Bartonella species C-values>1 (suggesting antibody production by the CNS) were detected in CSF of 11 cats, and B henselae DNA was amplified from the CSF of 10 cats. While the clinical significance of these findings cannot be assessed without a control group, the development of neurological signs in some cats inoculated with B henselae and the results of this study warrant prospective evaluation of the association of Bartonella species with feline CNS disease.     

Association of Trypanosoma vivax in extracellular sites with central nervous system lesions and changes in cerebrospinal fluid in experimentally infected goats

ABSTRACT: Changes in cerebrospinal fluid (CSF) and anatomical and histopathological central nervous system (CNS) lesions were evaluated, and the presence of Trypanosoma vivax in CNS tissues was investigated through PCR. Twelve adult male goats were divided into three groups (G): G1, infected with T. vivax and evaluated during the acute phase; G2, infected goats evaluated during the chronic phase; and G3, consisting of non-infected goats. Each goat from G1 and G2 was infected with 1.25 × 105 trypomastigotes. Cerebrospinal fluid (CSF) analysis and investigation of T. vivax was performed at the 15th day post-infection (dpi) in G1 goats and on the fifth day after the manifestation of nervous system infection signs in G2 goats. All goats were necropsied, and CNS fragments from G1 and G2 goats were evaluated by PCR for the determination of T. vivax. Hyperthermia, anemia and parasitemia were observed from the fifth dpi for G1 and G2, with the highest parasitemia peak between the seventh and 21st dpi. Nervous system infection signs were observed in three G2 goats between the 30th and 35th dpi. CSF analysis revealed the presence of T. vivax for G2. Meningitis and meningoencephalitis were diagnosed in G2. PCR were positive for T. vivax in all the samples tested. In conclusion, T. vivax may reach the nervous tissue resulting in immune response from the host, which is the cause of progressive clinical and pathological manifestations of the CNS in experimentally infected goats.     

Choroid plexus carcinoma cells in the cerebrospinal fluid of a Staffordshire Bull Terrier

An 11-year-old female intact Staffordshire Bull Terrier was referred to the Queen's Veterinary School Hospital at the University of Cambridge with sudden onset of episodic behavioral changes, a mammary mass, and papilledema in the right eye. On physical examination the dog appeared depressed and had a head tilt to the right with anisocoria. Using magnetic resonance imaging, a broad-based lesion that obliterated the fourth ventricle was detected in the right brainstem. There was no evidence of pulmonary metastasis. Cerebrospinal fluid (CSF) was then obtained; fluid analysis showed an increased cell count (165 cells/μL, reference interval 0-7 cells/μL) and total protein (0.30 g/L, reference value <0.25 g/L). Cytologic evaluation revealed a population of atypical epithelial cells arranged in cohesive rafts and characterized by moderate to occasionally marked anisocytosis and anisokaryosis. The appearance was highly suspicious of a malignant epithelial neoplasm. The dog was euthanized and on postmortem examination an asymmetrical nonencapsulated cerebellar mass was found within the choroid plexus of the fourth ventricle with local extension into the cerebellopontine angle. Histologic sections of the cerebellar mass contained arborizing papillary structures covered by a single layer of atypical epithelial cells that showed local infiltration into the adjacent neuropil. The diagnosis was choroid plexus carcinoma. The atypical epithelial cells were negative for pancytokeratin and strongly positive for vimentin. The finding of clusters of choroid plexus epithelial cells in the CSF demonstrates the value of utilizing a relatively noninvasive diagnostic technique for diagnosis of choroid plexus tumors.